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The multigene GAS family of Saccharomyces cerevisiae is constituted by five genes encoding GPI-anchored proteins required for cell wall or spore wall assembly. GAS1 and GAS5 are expressed in vegetative growth and repressed during sporulation, whereas GAS2 and GAS4 exhibit the opposite expression pattern. This study focuses on GAS3, a still poorly characterized member of the family. To date, attempts to reveal the glucan elongase activity typical of Gas proteins have been unsuccessful, suggesting that Gas3p is the only inactive member of the family. Here, we compared the mRNA levels of GAS1, GAS3 and GAS5 and demonstrate that GAS3 is the weakest-expressed paralogue in vegetative growth. Moreover, GAS3 mRNA increased during sporulation, showing a bimodal profile typical of the early-middle meiotic genes. GAS3 product was identified as a low-abundance, polydisperse mannoprotein. Loss of Gas3p did not affect growth and sporulation. The overexpression of GAS3, driven by the GAS1 promoter, slightly reduced growth rate in a wild-type strain and led to hyperaccumulation of Gas3p in the membranes and in the cell wall. To determine whether GAS3 could replace GAS1 function in vivo, GAS3 was also overexpressed in a gas1Δ mutant. Increased amounts of Gas3p were not only unable to complement the defects of the gas1Δ cells but exacerbated them. A mutated Gas3p-E283Q, where one of the catalytic glutamate residues essential for GH72 enzyme activity was replaced by glutamine, was also noxious to gas1Δ cells, indicating that the increased expression of Gas3p, rather than a potential activity, is deleterious for gas1Δ cells. Copyright © 2010 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The cell wall is the first interface between a fungus and its extracellular environment. Glycosyltransferases involved in the formation and dynamic remodelling of the polysaccharide network of the cell wall have recently been identified. The best characterized ones belong to the Gas family, which elongates β(1,3)-glucans, and to the Crh family, which are involved in the cross-linking of chitin to β(1,6)-glucan. All these proteins carry a glycosylphosphatidylinositol (GPI) anchor. In this work, we show that recombinant soluble forms of Gas1-5 and Crh1p from Saccharomyces cerevisiae and their orthologous proteins Gel1-Gel2 and Crf1 from Aspergillus fumigatus are specifically recognized by antibodies present in the sera of patients with Aspergillus or Candida infections. Quantification of the antibody titres against recombinant Gas/Gel and Crh/Crf proteins separated aspergilloma and candidiasis patients from non-infected individuals. Cross-reactivity was seen between the antibody response of patients with aspergillosis and candidiasis towards the Gas/Gel and Crh/Crf proteins. These results suggest that GPI-anchored cross-linking enzymes are relevant immunologically reactive constituents of the cell wall that may play a role during human fungal infections. Copyright © 2007 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

In budding yeast, chitin is found in three locations: at the primary septum, largely in free form, at the mother-bud neck, partially linked to β(1-3)glucan, and in the lateral wall, attached in part to β(1-6)glucan. By using a recently developed strategy for the study of cell wall cross-links, we have found that chitin linked to β(1-6)glucan is diminished in mutants of the CRH1 or the CRH2/UTR2 gene and completely absent in a double mutant. This indicates that Crh1p and Crh2p, homologues of glycosyltransferases, ferry chitin chains from chitin synthase III to β(1-6)glucan. Deletion of CRH1 and/or CRH2 aggravated the defects of fks1Δ and gas1Δ mutants, which are impaired in cell wall synthesis. A temperature shift from 30°C to 38°C increased the proportion of chitin attached to β(1-6)glucan. The expression of CRH1, but not that of CRH2, was also higher at 38°C in a manner dependent on the cell integrity pathway. Furthermore, the localization of both Crh1p and Crh2p at the cell cortex, the area where the chitin–β(1-6)glucan complex is found, was greatly enhanced at 38°C. Crh1p and Crh2p are the first proteins directly implicated in the formation of cross-links between cell wall components in fungi. [ABSTRACT FROM AUTHOR]